We have shown that antigen-induced release of secretory granules from a rat mast cell (RBL-2H3) line is dependent protein kinase C whereas release of arachidonic acid is dependent on MAP kinase for the phosphorylation and activation of phospholipase (PL) A2. Both processes are dependent on an increase in calcium for activation of secretory machinery and translocation of the phosphorylated PLA2 to membrane and, as shown here, recruitment of the tyrosine kinases, Lyn and Syk. Further elaboration of this scenario stems from work in three areas. With respect to protein kinase C, the activating diglycerides may be generated via PLD as well as PLC as indicated by stimulation of cells through adenosine A3 receptors. This stimulation results in transient activation of PLC but persistent activation of PLD and protein kinase C. With respect to MAP kinase, the entire MAP kinase/PLA2 cascade (i.e. the sequential phosphorylation of Ras/Raf->MEK-1->p42mapk->cPLA2) is activated by several mechanisms which include Syk (possibly via Grb2, Sos and Shc), elevation of free Ca2+, activation of protein kinase C or engagement of trimeric proteins. Each of these mechanisms can be selectively activated or inhibited and their ability to activate the MAP kinase/PLA2 cascade can be blocked by dexamethasone which may act at a step immediately prior to the interaction of Ras with Raf. These and other studies indicate discrete pathways for activation of the cascade which converge at the level of Ras/Raf. Finally, with respect to the secretory machinery, RBL-2H3 cells acquire sensitivity to the mast cell secretagogue, compound 48/80, under conditions that are thought to induce phenotypic alterations in the distribution of components that participate at late steps of exocytosis. Compound 48/80 has been previously shown to induce secretion in rat peritoneal mast cells by directly activating G(i-3) in the plasma membrane. Changes in distribution of this G protein, which is normally confined to intracellular organelles in RBL-2H3 cells, and other candidates are under study.